Drosophila yakuba is widespread in Africa. Here we compare the cuticular hydrocarbon (CHC) profiles and mating behavior of mainland  and island  populations. The strains each had different CHC profiles: Bioko and Kounden were the most similar, while Mayotte and Sao-Tome contained significant amounts of 7-heptacosene. The CHC profile of the Sao-Tome population differed the most, with half the 7-tricosene of the other populations and more 7-heptacosene and 7-nonacosene. We also studied the characteristics of the mating behavior of the four strains: copulation duration was similar but latency times were higher in Mayotte and Sao-Tome populations. We found partial reproductive isolation between populations, especially in male-choice experiments with Sao-Tome females. Drosophila cuticle. They protect against desiccation and stimulate courtship [melanogaster subgroup, and there is evidence that CHCs with at least one double bond in position (Z)-7 function as pheromones [Long-chain cuticular hydrocarbons (CHCs) are fatty acid-derived hydrocarbons present on the ourtship ,2. CHCs eromones .melanogaster subgroup the chemical composition of CHCs is quite similar in males regardless of the species, with very abundant monoenes in C23 and C25 (7-tricosene (7-T) and 7-pentacosene (7-P)), except in D. erecta [D. melanogaster, D. sechellia and D. erecta. The remaining six species have similar CHC profiles in males and females: D. yakuba, D. santomea, D. teissieri, D. orena, D. mauritiana and D. simulans [In the . erecta . CHC comsimulans .D. melanogaster and D. simulans [D. sechellia and D. mauritiana [D. sechellia and D. simulans [D. melanogaster, a female CHC polymorphism is linked to the geographic origin of two populations, characterized by 5,9-dienes in African and Caribbean populations and 7,11-dienes in all other populations [D. santomea and D. yakuba [D. melanogaster [D. simulans [Female pheromones contribute to reproductive isolation between simulans , D. sechuritiana , and D. simulans . Within ulations . This CH. yakuba , and betsimulans .melanogaster subgroup species originated in the African tropics. Some of them are cosmopolitan , some are island endemics , or more or less confined to a restricted area of the African mainland . One (D. erecta) is specialized on Pandanus fruits. The last species (D. yakuba) is broadly distributed across tropical sub-Saharan Africa [All the n Africa . It is an Africa .D. yakuba might show some geographic differentiation, leading to reproductive isolation between different populations. To address this question, we performed pheromone analyses and mating tests on one population from mainland Cameroon and three populations confined to different African islands.Because of its widespread distribution, we wondered whether D. yakuba flies were maintained on standard yeast/cornmeal/agar medium in a 12-h-light/12-h-dark cycle at 25 °C. We used four strains, issued from isofemale lines (given and collected by J. David in 2013). One was from the mainland in Cameroon: Kounden (K) . Three were from different islands: Bioko (B) ; Sao-Tome (S) from the west coast of Africa ; and Mayotte (M) from southeast Africa . All strains were maintained in the laboratory for two years. Flies were sexed, isolated under carbon dioxide anesthesia within 2 h of imaginal emergence, and kept in sex-specific groups of 10 in fresh food vials until testing. Five-day-old flies were used for the experiments.n = 10 for all measurements).CHCs were extracted from individual 5-day-old flies by washing them for 5 min in 100 µL of heptane containing 500 ng hexacosane (C26) as an internal standard. The fly was then removed from the vial and 5 µL of each sample was injected into a Perichrom Pr200 gas chromatograph with hydrogen as the carrier gas, using a split injector (split ratio 40:1). The column was a 25 m long BP-1 that had an internal diameter of 220 μm and 0.1 μm film thickness (SGE). The oven temperature started at 180 °C then ramped at 3 °C/min to 300 °C, for a total run of 40 min. The flame ionization detector was set at 260 °C . CHCs wen ≥ 50 for all tests).All tests were run in the morning, between 9 and 12 a.m. under dim electric light in a temperature-controlled room. For the mating tests, we used a watchglass of 28 mm diameter and 5 mm internal height glass placed it on a glass plate as an observation chamber. We introduced a female into the observation chamber and left her for 1 min before introducing a male. We recorded copulation latency  and the duration of copulation. Individuals that did not mate within 45 min were not used in the analysis of copulation latency  for choice tests, to allow the flies to access the food and to give females sufficient room to escape from males. We cut a small portion of one wing  of all the flies to allow us to differentiate flies of different strains in mate-choice tests. For female-choice and male-choice tests, a single female  was transferred to a fresh food vial together with two males  from different strains. The trial ended once the female had copulated with one of the males.For multiple-choice tests, two males and two females  were introduced into a fresh food vial. The trial ended when the first copulation occurred. We did the multiple-choice tests because they are more sensitive at detecting assortative mating than are no-choice tests and they allow both male and female choice to contribute to assortative mating . We carrAll statistical analyses were performed using R version 3.2.1 .CHC data were normalized relative to 2-methylhexacosane (Me26), which was proportionally the least variable compound. The analysis was thus performed on the logarithm of the ratio between measured abundance and Me26. We replaced measurements of 0.0 with 0.1 (close to the detection limit with our protocol) before log transformation.manova function in R), and p-values were corrected for multiple tests (Holm-Bonferroni correction). When the population factor appeared to be significant in the ANOVA, we used post-hoc analyses to compare pairwise differences among populations (Tukey HSD test). In addition, we used principal component analysis to evaluate abundances (function prcomp in R).The effects of sex, population, and the sex-by-population interaction were tested using a two-way multivariate ANOVA  to quantify departure from the random mating hypothesis . Multiple testing-corrected p-values (Holm-Bonferroni correction) are indicated as p*. The specific influence of male/female population on the homogeneous/heterogeneous cross ratio was investigated by running binomial generalized linear models (GLM):m and f are the population of the introduced male/female, µ is the reference probability , xm and yf describe the effects the male and female population and zmf is the interaction between both populations. The link function g was the default logit function. Conveniently, depending on the hypothesis being tested, the intercept could be set to µ = 0  or it could be computed arbitrarily as the average of the first experiment (in which case the model estimates differences between experiments). The general significance of the factors was assessed with an analysis of deviance (function anova.glm in R), which determines the most parsimonous models by adding factors one by one (forward model selection) and performing likelihood ratio tests. As the factor order matters in such models, we made sure to introduce the factors in the following order: (i) the population of the tested individual ; (ii) the population of the heterogeneous cross ; and (iii) the interaction between the two.In the 1 + 2 mating experiments , we considered three categories: Homogeneous crosses, heterogeneous crosses, and no mating. We used the The same setting was used to test the influence of male/female populations on the proportion of trials in which there were no mating events . Finally, the same factors were considered for the analysis of copulation latency (linear model after log-transformation).0A); (2) equal numbers of each kind of homogeneous crosses (H0B); and (3) random mating (H0C), in which 25% of the mating were homogeneous crosses of each type and 50% were heterogeneous crosses. For H0C, for convenience, the multinomial p-value was computed by Monte-Carlo simulations .In the 2 + 2 experiments, four categories were defined: homogeneous for population 1, homogeneous for population 2, heterogeneous, and no mating. For each experiment, we tested three hypotheses: (1) equal homogeneous and heterogeneous crosses . There was also a large discrepancy in the amount of C23 . Sao-Tome females had the most C25, C27 and C29 .Bioko males more often exhibited courtship and eventually mated than did males from the other populations. Around 50%, of Kounden, Mayotte and Sao-Tome males mated—about 28% less than Bioko . Latencyvs. 177 heterogeneous, p-value < 0.001 in the 1 female/2 male experiment; 219 vs. 145 pairs, p-value < 0.001 in the 1 male/2 female experiment; and 126 vs. 67 pairs, p-value < 0.001 in the 2 males/2 females experiment)  .p = 0.04), but no effect of the male population (p = 0.32). The GLM with no intercept (test for the departure from random mating) attributes this effect to females from the Mayotte population (p = 0.01) and from the Sao-Tome population (p < 0.001), while Bioko and Kounden females seem to mate randomly . There was also a significant female effect on the proportion of non-mating events . Females from populations Bioko and Kounden mated more often than females from populations Mayotte and Sao-Tome. We found no significant effect from population on copulation latency for either females (p = 0.06) or males (p = 0.64).In the female choice experiment , the anap = 0.13 in the analysis of deviance) or female (p = 0.16) population on the proportion of homogeneous crosses, although some specific crosses  departed significantly from random mating hypothesis on their own. In contrast, there was a significant effect of female population on the proportion of pairs in which there was no mating (p = 0.009)—an effect that was not observed for males (p = 0.11). More specifically, the GLM identified a significant departure of females from the Sao-Tome population (p = 0.0028) compared to the Bioko population, which was taken arbitrarily as a reference. This strongly suggests that the presence of a Sao-Tome female in the experiment prevented any male from another population from mating with the other female. We found no significant effect of male (p = 0.46) or female (p = 0.05) population on copulation latency.In the male choice experiment, we found no evidence of a differential effect of male (p = 0.002). In contrast, Sao-Tome and Bioko populations were the only ones driving a significant deviation from an even representation of both kinds of homogeneous pairs . Sao-Tome population showed a general trend towards being preferentially associated with such deviations, but this average effect was not significant (p = 0.057). We did not detect any population effect on copulation latency or on the proportion of pairs in which there were no mating events.In the multiple-choice test, testing for random mating is more complicated as there can be deviations from the 1:1 ratio of homogeneous/heterogeneous crosses and/or uneven frequencies of homogeneous crosses. A binomial GLM with no intercept showed that only the Mayotte population had a significant influence on the excess of homogeneous crosses  [D. yakuba is also found at low elevation. In Sao-Tome, the D. yakuba strains occurring at high altitude are thought to be more ancient that the ones occurring at low altitude, because D. santomea, which diverged from a common ancestor with D. yakuba 400,000 years ago [D. yakuba throughout lowland Sao-Tome may be the result of a recent secondary invasion [D. yakuba strain collected at low altitude, showed a CHC profile similar to that of Bioko and Kounden. These results strengthen the hypothesis that D. yakuba flies at lowland sites were recently introduced.The Sao-Tome strain used in this study was collected at about 1200 m altitude in a zone where hybrids between cy (~1%) ,20. D. years ago ,22, inhainvasion . Mas andinvasion , analysiD. melanogaster (about 20 min) under the same conditions. An even longer copulation duration has been reported in D. yakuba (60 min for the Abidjan strain). Copulation in most Drosophila species lasts about 30 min [Copulation latencies were high for all strains, compared to others reported in the literature ,23 , courtship occurs under five seconds and mating within the first two minutes both in the laboratory and the field. In this case, no-choice designs are reflecting nature better than choice-designs [D. yakuba and in these conditions, choice-designs seem more appropriate than no-choice ones because females have the time to choose a mate between different males. However, we cannot prevent any bias resulting from the different copulation latencies within D. yakuba populations. We have used one-choice and multiple-choice experiments. Although some authors suggest that multiple-choice designs are more realistic because flies congregate before mating on food, this does not seem to be justified, because very little is known on the circumstances of mating, the density and sex-ratio of flies in the mating habitat [Although the proportion of homogeneous matings was generally higher than the proportion of heterogeneous ones, we lacked statistical power to demonstrate significant reproductive isolation between strains, except between Kounden and Mayotte (multiple-choice tests). In previous unpublished experiments, we measured mate choice using the small watchglass apparatus used here to measure courtship behaviour, and observed strong reproductive isolation between strains. We cannot totally explain this discrepancy. In nature, flies have continual access to food, which has an attractive role and is also important for mating behavior. The device used in this study is more appropriate for choice experiments because flies can smell food, eat, fly and escape from an undesired partner. In some species  [D. yakuba populations. The rapid genome evolution described in isogenic lines from D. yakuba [We were able to demonstrate a peculiar mating behavior within the Sao-Tome population. As CHC profiles differ between the strains, it is tempting to emphasize the role of CHCs in mate choice. Although CHCs contribute to reproductive isolation in most species within the melanogaster subgroup, other factors, such as courtship songs, could play a meaningful role . The melic songs . Sexual  females . D. santhip song . D. sant(rowing) . The evo. yakuba  might dr


Copro-parasitological diagnosis is still a challenge in management of helminth infections at individual and community levels in resource-limited settings.The aim of our study was to compare the performance of three quantitative techniques: Kato-Katz, McMaster and Mini-FLOTAC methids. The study was carried out in Oran, Northern Argentina.200 schoolchildren were enrolled to provide a single stool sample, which was tested for helminth infections with Kato-Katz, McMaster and Mini-FLOTAC methods. The Mini-FLOTAC was performed with two flotation solutions . Preparation and reading time for each of the three methods was calculated both when processing single and multiple samples.Hymenolepis nana (23%) followed by Ascaris lumbricoides (17%) and a third group of less prevalent helminths: Enterobius vermicularis, Trichuris trichiura and hookworms . Mini-FLOTAC FS2 was more sensitive than FS7 for H. nana (93% vs 78%) and for other helminths (85% vs 80%), whereas FS7 was more sensitive for A. lumbricoides (87% vs 61%). Kato-Katz method was more sensitive than McMaster method for A. lumbricoides (84% vs 48%) and for other helminths (48% vs 43%) except for H. nana (49% vs 61%). As for egg counts, Mini-FLOTAC FS2 reported 904 eggs per gram of faeces (EPG) for H. nana (vs 457 with McMaster and 111 with Kato-Katz) and 1177 EPG for A. lumbricoides (vs 1315 with Kato-Katz and 995 with McMaster); FS2 detected the highest EPG for both H.nana and A.lumbricoides (904 vs 568 and 1177 vs 643 respectively), the differences were not statistically significant. The technique feasibility was calculated: Kato-Katz mean time was 48 minutes/sample, Mini-FLOTAC 13 minutes/sample and McMaster 7 minutes/sample. However, especially for Kato-Katz and Mini-FLOTAC, the mean time (min/sample) decreased significantly when processing multiple samples.Out of 193 schoolchildren examined, 40% were positive for any helminth infection by any method; the most prevalent was H. nana and as sensitive as Kato-Katz and more sensitive than McMaster for A. lumbricoides identification. Egg counts differences although relevant, did not reach statistical significance.Mini-FLOTAC is a promising technique for helminth diagnosis, it is more sensitive than Kato-Katz and McMaster for  Ascaris lumbricoides, Trichuris trichiura, Ancylostoma duodenale/Necator americanus) infections still represent a great burden in public health and affect almost two billion people worldwide[Soil-transmitted helminths (STH)  and Strongyloides stercoralis (24%). These data were confirmed in a recent review by Socias et al.[In 2007 Menghi et al. conductes et al.. These rKato-Katz is the quantitative technique recommended for the diagnosis of STH. McMasteIn addition to comparing diagnostic accuracy, this study also focused on the affordability of this innovative technique and on its transferability to peripheral laboratories, in order to improve diagnosis and facilitate parasitological monitoring of STH control programmes.San Ramon de la Nueva Oran is located in Northern Argentina, Salta Province, 32 km south from the Bolivian border. It is surrounded by rivers and therefore agriculture is the main economic resource for the population. The last National census in 2010 registered approximately 67,000 inhabitants. The climate is subtropical, characterized by a dry (April-October) and a humid (November- March) season. This survey was carried out in August-November 2012, during late winter and spring.The Laboratory of Parasitology and Tropical Medicine is situated at the local branch of the National University of Salta; specimens from the primary care service are directed there for parasitological examinations free of charge.This study was carried out examining stool samples from one of the primary schools of the town. The school was randomly chosen among the schools in the suburbs of Oran where STH were thought to be more widespread. All children in the school were examined for STH infections to reach 200 children. This number was calculated, assuming a prevalence of STH of about 50% from previous studies, in order to have at least 100 positive children, to allow a power of 80% and a difference of 20% between the diagnostic methods with a 95% confidence level-13.All stool samples were collected and analyzed within 24 hours with the three different techniques: Kato-Katz, McMaster and Mini–FLOTAC.The Kato-Katz was performed using the 41.7 mg template, according to WHO recommendations.McMaster method was performed as the standard procedure: 2 g of faeces were filtered and homogenized with 30 ml of saturated saline. Two flotation chambers (1 ml each) were filled for each sample and three minutes were needed for the eggs to float. Eggs were counted and multiplied by 50 to have the eggs per gram of faeces (EPG).Mini-FLOTAC is a new diagnostic method based on flotation of the eggs. Two chambers (1 ml each) are placed in the base and surmounted by a reading disc. The Mini-FLOTAC is part of a kit together with the fill-FLOTAC, a plastic device used to homogenize, filter and pour the sample in the flotation chambers. The floA. lumbricoides was considered according to WHO guidelines: light infection <5000 EPG, moderate 5000–49 999 EPG and heavy ≥50 000 EPG[Intensity of infection for  000 EPG.The feasibility of the 3 techniques was evaluated on a total number of 100 samples randomly assigned to 4 experienced laboratory technicians. The time needed to prepare the samples and to clean the devices was measured six times. The preparation period started when the samples were weighed and ended when the sample was ready for examination. During the cleaning period all used materials were either disposed, for single use components, or cleaned in the case of recyclable devices. Examination of the slides or chambers was timed individually for all samples. Each sample was examined with all techniques by the same laboratory technician.Data were analyzed by EPIDAT version 3.1 . Prevalence, sensitivity (Se), negative predictive value (NPV) and Kappa index (KI) for each method  were calculated. Prevalence was defined as proportion of positive samples over total samples analyzed. A positive sample was defined if positive with any parasitological method, while a negative sample was considered negative if negative with all methods; this criterion was defined our “gold standard”. Sensitivity  and NPV (proportion of negative results among un-infected subjects) were calculated using 2×2 contingency tables in relation to our “gold standard”. The agreement between the results of each method was calculated with the KI. Interpretation of KI was as follows: ≤0 = poor, 0.01-0.20 = slight, 0.21-0.40 = fair, 0.41-0.60 = moderate, 0.61-0.80 substantial, and 0.81-1.00 almost perfect agreement.The study was reviewed and approved by the Ethics Committee of the Faculty of Medicine, San Raffaele Hospital, Milan, Italy. A separate ethical clearance was obtained from the San Ramon de la Nueva Oran Hospital Management Board. The medical staff met school authorities and parents/guardians before enrolling their children in the survey and verbal informed consent was sought. After the survey, all schoolchildren were treated with mebendazole 500 mg in single dose according to the WHO recommendations for preventive chemotherapy. Data weHymenolepis nana, 32 (16.6%) were positive for Ascaris lumbricoides, few others were found positive for other helminths: Enterobius vermicularis, Trichuris trichiura and hookworms . For A. lumbricoides only two heavy infections were detected, nine were moderate and twenty-one light.Out of two hundred children who were enrolled in the survey, 193 provided the stool samples. The enrolled children aged between 5 and 13 years old (mean 10 years), and 82 (42.5%) were males; 78 children (40%) were found positive for at least one helminth infection. Forty five (23%) children were found positive for H.nana was Mini-FLOTAC FS2 (92.7%) followed by FS7 (77.8%), McMaster (60.5%) and Kato-Katz (49.0%); for A.lumbricoides the most sensitive was Mini-FLOTAC FS7 (87.1%), followed by Kato-Katz (84.4%), Mini-FLOTAC FS2 (61.3%) and McMaster (48.3%). The difference in helminth detection among the three techniques was statistically significant. Mini-FLOTAC FS7 and Kato-Katz method detected significantly more infections than McMaster (p < 0.05) for A. lumbricoides diagnosis. Mini-FLOTAC FS2 was more sensitive than both McMaster and Kato-Katz for H.nana (p < 0.05). Regarding the two flotation solutions, FS2 was more sensitive than FS7 for H.nana (92.7% vs 77.8%), but the latter was more sensitive for A.lumbricoides (87.1% vs 61.3%), although the differences were not statistically significant. The performance of the test on negative subjects (NPV) was high: > 85% for H. nana and > 90% for A lumbricoides with all methods. NPV for H.nana with Mini-FLOTAC was significantly higher than with Kato-Katz and McMaster method. However, NPV is greatly influenced by the prevalence of infection and its results may vary in other settings with different STH distribution. The prevalence of the other helminths was not high enough for statistical analyses.Each sample was analysed with the three different techniques and results are shown in Figure H.nana between Kato-Katz thick smear and Mini-FLOTAC FS7 and McMaster method (0.57 and 0.58 respectively); it was substantial between Kato-Katz and FS2 (0.63), McMaster and Mini-FLOTAC FS2 and FS7  and between Mini-FLOTAC FS2 and FS7 (0.85). For A.lumbricoides infection the agreement was moderate between Kato-Katz and McMaster (0.59) and between McMaster and Mini-FLOTAC FS7 (0.57), was substantial between Kato-Katz and Mini-FLOTAC FS2 (0.78), McMaster and Mini-FLOTAC FS2 (0.67) and between Mini-FLOTAC FS2 and FS7 (0.76); it was almost perfect between Kato-Katz and Mini-FLOTAC FS7 (0.98).The KI for agreement among techniques is shown in Table H.nana (904), followed by Mini-FLOTAC FS2, McMaster and Kato-Katz . For A.lumbricoides Kato-Katz detected the highest number of EPG (1315) followed by Mini-FLOTAC FS2 (1177 EPG), McMaster (995 EPG) and Mini-FLOTAC FS7 (643 EPG). Due to the large variance, the EPG values were not statistically different among the three methods.As the three techniques are quantitative diagnostic methods, comparison among EPG was also calculated and data are shown in Table A.lumbricoides light infections were the most frequent to be missed, in fact 12 negative samples at the McMaster resulted positive for light infections with the Kato-Katz, 7 negative with the Kato-Katz resulted positive for light infections with the Mini-FLOTAC FS2, and 6 negative with McMaster were positive for light infections with Mini-FLOTAC FS2; between Mini-FLOTAC FS2 and Mini-FLOTAC FS7, 9 negative with the former solution were positive for light infections with the latter.As shown in Additional fileOverall, the most time-consuming method to be processed was Kato-Katz (mean time 48 min/sample), followed by the Mini-FLOTAC (13 min/sample). McMaster was the quickest to process (7 min/sample). Kato-Katz was the slowest to examine (10 min/sample), followed by the Mini-FLOTAC technique with FS7 (7 min/sample). The McMaster and Mini-FLOTAC with FS2 slides were the quickest to read (5 min/sample).The waiting time for the clarification of the eggs for Kato-Katz and for the floatation of the eggs for the Mini-FLOTAC were the factors that accounted most significantly for the length of processing single samples. It must be underlined, however, that for both techniques, when processing multiple, samples the time (min/slide) decreased significantly as the reading of the slide(s) was done while awaiting the cleansing of subsequent samples. Reading time was faster for the McMaster and Mini-FLOTAC FS2 as they were clearer slides, whilst Mini-FLOTAC FS7 and the Kato-Katz were slides more difficult to read due to the presence of debris and artefacts. Examples of slides with helminth eggs with the three different methods are illustrated in Figure A. lumbricoides as the prevalence of the other two major STH, T. trichiura and hookworms, was too low for a meaningful evaluation. As for the comparison with McMaster method, Mini-FLOTAC proved to be more sensitive for all helminths, especially for A. lumbricoides and H. nana. This is the first study that reports the detection of H. nana with the Mini-FLOTAC and was shown to be a sensitive method for this particular parasite. Mini-FLOTAC also counted more eggs than Kato-Katz, which is not the best technique to assess intensity of infection of H. nana. One possible explanation could be that H. nana eggs are very small (30–40 μm) and yellowish/transparent and therefore could not easily be seen in the Kato-Katz. Rather they are well detected upon flotation in the Mini-FLOTAC. Even if H. nana is not included in helminth control programmes, studies conducted showed its pathogenicity and occasionally severe symptoms linked to this infection. It would be interesting, in the light of this reported data, to further investigate H.nana epidemiology, perhaps using the Mini-FLOTAC, which was demonstrated to be a sensitive diagnostic method for its detection[H. nana and is more sensitive than the McMaster for A. lumbricoides. Advantages of the Mini-FLOTAC are its relatively quick processing and read out and its good sensitivity which compete with the McMaster (simpler but less sensitive) and with the Kato-Katz (as sensitive but more time-consuming).The Mini-FLOTAC turned out to be an innovative, sensitive and low cost technique in the scenario of intestinal helminths diagnosis; it has been recently launched, so some aspects are still on trial, but recent studies revealed a potentially reliable diagnostic tool,16. Minietection-19. The Another added value of the Mini-FLOTAC is that it works in a “closed” system (using Fill-FLOTAC) and that faeces can be preserved with 5% formalin in order to guarantee the safety of the operator as well as the possibility of examining the samples up to one week after their collection. Regarding floatation solutions, FS7 is less easily available as zinc sulphate is not cheap and relatively difficult to find in low-resources settings, whilst FS2, which is based on saturated salt solution, is cheap and easily accessible. Regarding feasibility, when multiple faecal samples are examined in series, the processing time and reading differences of the three methods flatten, and are more or less similar among the three techniques, with a slight advantage for the McMaster. Among all determinants influencing the length of each technique, faecal egg count was the most relevant, especially for Kato-Katz.In situations where helminth control strategies aim at control of transmission and eventually elimination of STH, the need of a diagnostic tool easy to use in the field and able to detect even very light infections is evident. The need of new sensitive diagnostic methods has been partially met by the introduction of the FLOTAC technique for direct diagnosis, and molThe major limitation of this study is the low prevalence of helminth infections which hinders the full evaluation of the Mini-FLOTAC against a wide range of intestinal parasites. It would be useful, therefore, to plan other studies to compare these three techniques in areas where the prevalence of other STH is higher. A higher prevalence of STH in this area, especially hookworm was reported-6,13 a fThe ideal direct diagnostic method in helminthology should combine ease of use, affordability, and relatively good sensitivity, especially in quantitating egg counts. The data presented show that all three techniques tested in this study match these criteria, with a slight advantage for the Mini-FLOTAC with FS2. The techniques evaluated in this study should be recommended for STH mapping purposes, monitoring drug efficacy and impact of STH control programme interventions in endemic countries.Mini-FLOTAC and Fill-FLOTAC were invented and patented by Prof. Giuseppe Cringoli, University of Naples Federico II, Italy. However, there was no conflict of interest in the study.BB, MA, RB, GC and AK designed the project. RC, LR made statistical analysis. BB, PC, EV, MJ carried out the work field and laboratory analyses. GC, RB financially supported the project. BB, MA, AL wrote the paper. All co-authors gave important contributions in revising the manuscript and answering the reviewers. All authors read and approved the final version of the manuscript.2×2 contingency table of A. lumbricoides intensity of infections with the three diagnostic methods.Click here for file


Fearless removal of tooth structure during canal preparation and shaping has negative effects on the prognosis of treatment. On the other hand, sufficient pre-enlargement facilitates exact measurement of the apical size. The present A total of 40 mandibular molars were selected and the coronal areas of their mesiobuccal and mesiolingual root canals were randomly prepared with either mentioned instruments. Pre- and post-instrumentation CBCT images were taken and the thickness of canal walls was measured in 1.5- and 3-mm distances from the furcation area. Data were analyzed using the one-way ANOVA. Tukey’s post hoc tests were used for two-by-two comparisons. P<0.02); however, two-by-two comparisons did not reveal any significant differences between the other groups. At 1.5-mm distance, there was no significant difference between different instruments. However, at 3-mm distances, Gates-Glidden drills removed significantly more dentin compared to FlexMaster files (mean=0.18 mm) ( All tested instruments can be effectively used in clinical settings for coronal pre-enlargement. Increasing use of engine-driven instruments during root canal preparation, necessitates a correct understanding of their properties and limitations. NiTi instruments are superior to stainless steel files due to the superelastic properties. This characteristic make endodontic files more elastic and increase their compliance with the root curvature and their resistance against fracture . The aim of cervical preparation is to gain direct access to the apical area of the canals or the apical curvatures . The ceraka the danger zone) have thinner distal walls which might be endangered during the use of different instruments for endodontic treatment. Recently, Mahran and AboEl-Fotouh [et al. [The mesial roots of mandibular molars and the mesiobuccal roots of maxillary molars  and three finishing files  [FlexMaster files  have a triangular cross-section with K-type cutting blades and no radial lands. Different tip sizes are also available with 2, 4 and 6% tapers. IntroFile (20/0.11) is the primary orifice shaper with 11% taper and a 9-mm cutting blade. ProTaper system  is amongst the pioneer engine driven instruments with full 360RaCe instruments (Reamer with Alternating Cutting Edges)  have a triangular cross-sectional design with alternative cutting edge which is aimed at reducing the tendency to thread the file into the root canal. The exception is the 20/0.02 files, which have a square cross-section. RaCe is marketed with variable tapers  , 11, 12.radial land relief, which aids in protecting the file from over engagement in the canal. The K3 system also offers a third radial land to help prevent threading. The instruments are available with 12, 10, 8, 6 and 4% tapers [K3 instruments  have K3 instruments  have a slightly positive rake angel for greater cutting efficiency, wide radial land  and feature a % tapers -16.CBCT is an imaging system which provides three-dimensional (3D) scans from the maxillofacial skeleton; it has overcome the limitations of intraoral radiographic techniques. Compared to the conventional intraoral radiographic techniques, CBCT is more effective in collecting sufficient information for the diagnosis and achieving more capabilities in the management of complicated problems of endodontics , 17. in vitro study was to compare the efficacy of all aforementioned systems  in coronal pre-enlargement of root canals, using CBCT.The aim of the present °C for disinfection. The teeth were washed with running tap water 24 h before use, to eliminate traces of thymol and were then stored in normal saline at 4°C until further processing. Assessment radiographic images were taken using E-speed films  with 70 kVp and 8 mA; the films were processed by a Hope film processor. The exclusion criteria included a more-than-3 mm distance between the CEJ and furcation area on radiographs, previous endodontic treatment, incomplete root formation, signs of internal root calcification, external or internal root resorption and more than 40° root curvature . All thGroup 1 (Gates-Glidden drills): The root canals were prepared using #3, 2 and 1 Gates-Glidden drills  installed on a low-speed handpiece operating at 12.000 rpm. The drills were used directionally in an anti-curvature mode to selectively remove dentin from the bulky wall (safety zone) toward the line angle, protecting the danger zone; Group 2 (K3): In this group, 25/0.12, 25/0.10 and 25/0.08 files were used with a gear reduction handpiece powered by a on an electric motor  set at a speed of 300 rpm and torque of 2 Nm; Group 3 (ProTaper): Root canal preparation was carried out with SX, S1 and S2 instruments set on the same device with speed and torque of 300 rpm and 3 Nm, respectively; Group 4 (FlexMaster): IntroFile (20/0.11) was used for coronal pre-enlargement with a speed of 300 rpm; Group 5. (RaCe): In this group, 40/0.10 and 35/0.35 files were used with a speed of 600 rpm and the torque was set at 1.5 Nm. The working length (WL) was determined for preparation of the canals. A #10 K file  was placed in the root canal so that its tip was visible at the apical foramen; the WL was set 1 mm short of the file length. The teeth were randomly divided into 5 groups including 8 mesiobuccal and 8 mesiolingual root canals in each group. The groups were instrumented as follows: The root canals were irrigated with 2 mL of 2.5% NaOCl between instruments. After preparation, the root canals were irrigated with saline and 2 mL of 2.5% NaOCl to remove all dentin debris. Each file series was discarded after use in one canal or when any defect or deformation was observed in the file. Debridement was carried out by one operator. The operator debrided the canals at a specific time of the day and worked only on 5 canals each day so that a constant and uniform force would be applied during canal preparation and the operator fatigue would not exert any effect on the results. Then the samples were placed in the CBCT unit and a post-instrumentation image was taken in the same manner. Then, 0.5-mm axial cross-sections were prepared at 1-mm intervals. The mean values of dentine removal, standard deviations, mean standard errors and 95% confidence intervals of interval differences of the amounts of dentin removed were calculated before and after preparation with different instruments and the data was analyzed with one-way ANOVA. Due to the presence of statistical significance at 3-mm cross-sections, Tukey’s post hoc test was used for two-by-two comparisons of instruments in relation to the amount of dentin removal.P=0.93). The mean±SD of the dentin removal value at 1.5- and 3-mm distances from the furcation, were 0.280±0.22 and 0.278±0.22 mm, respectively with no statistically significant differences (P=0.06); however, the differences at 3-mm cross-sections were significant (P<0.025). P<0.02); however, the differences between other groups were not significant. At 1.5-mm sections, two-by-two comparisons were not made because one-way ANOVA did not reveal any significant differences among the test groups.Two-by-two comparisons of instruments at 3-mm sections showed that differences in dentin removal were only significant between FlexMaster and Gates-Glidden instruments (in vitro study compared the coronal-enlargement efficacy of different endodontic instruments using CBCT. Nowadays use of CBCT imaging technique has gained attention due to easy access to processing programs such as Photoshop [et al. [et al. [et al. [This hotoshop . This tehotoshop  which ishotoshop -18. In ahotoshop . Hartman [et al. , showed  [et al. . In prev [et al. , histolo [et al. , electro [et al. , serial  [et al.  and radi [et al. , used muCBCT technique was used in the present study to evaluate the samples due to the advantages mentioned above. There were no significant differences between different root canal preparation systems and instruments in the amount of dentin removal in 1.5-mm cross-sections. However, at 3-mm apical to the furcation, Gates-Glidden drills removed significantly more dentin compared to FlexMaster files. Other two-by-two comparisons did not reveal any significant differences among different systems.et al. [et al. [et al. [Sanfelice et al.  did not et al. . However [et al.  were dif [et al. -27. Baseet al.[In the study by Flores et al., no diffet al. [In another study by Kássio et al. , no diffet al. [Marco et al.  comparedet al. [In the study by Sanfelice et al. , where nAll the tested instruments had similar efficacy in coronal pre-enlargement and are safe enough for clinical use.


Several approaches have been used to express energy expenditure in youth, but no consensus exists as to which best normalizes data for the wide range of ages and body sizes across a range of physical activities. This study examined several common metrics for expressing energy expenditure to determine whether one metric can be used for all healthy children. Such a metric could improve our ability to further advance the Compendium of Physical Activities for Youth.2) data obtained from five sites was completed, that included 947 children ages 5 to 18 years, who engaged in 14 different activities. Resting metabolic rate (RMR) was computed based on Schofield Equations . Absolute oxygen uptake (ml.min-1), oxygen uptake per kilogram body mass (VO2 in ml.kg-1.min-1), net oxygen uptake (VO2 – resting metabolic rate), allometric scaled oxygen uptake (VO2 in ml.kg-0.75.min-1) and YOUTH-MET (VO2.[resting VO2] -1) were calculated. These metrics were regressed with age, sex, height, and body mass.A secondary analysis of oxygen uptake (VO2, and YOUTH-MET were least associated with age, sex and physical characteristics. For moderate-to-vigorous intensity activities, allometric scaling was least related to age and sex. For sedentary and low-intensity activities, YOUTH-MET was least related to age and sex.Net and allometric-scaled VONo energy expenditure metric completely eliminated the influence of age, physical characteristics, and sex. The Adult MET consistently overestimated EE. YOUTH-MET was better for expressing energy expenditure for sedentary and light activities, whereas allometric scaling was better for moderate and vigorous intensity activities. From a practical perspective, The YOUTH-MET may be the more feasible metric for improving of the Compendium of Physical Activities for Youth. Physical activity (PA) plays an important role in health and in normal growth and development . SubstanThe youth compendium is comparable to that used in adults ,5 and ha.min-1), similar to oxygen uptake (VO2) per min (ml.min-1), increases with age as a function of body mass or muscle mass. Thus, it is not appropriate to compare absolute EE of children and adolescents of varying sizes. Several approaches have been used to normalize EE values for differences in body mass in order to compare children and adolescents; thus an effort to examine various metrics of youth energy expenditure during different physical activities across a range of ages is important for improving our capacity to compare EE using a common scale.Absolute EE  has been criticized by Nevill et al. [.kgFFM-1.min-1 [Approaches for normalizing EE for body mass can be classified into three categories: 1) ratio scaling of EE for body mass –15; 2) ul et al. ,17 becaul et al. ,7. Since-1.min-1 ,19. Howe-1.min-1 . Also, tResearchers have used allometric scaling to adjust EE for size-related changes in physiological functioning and the disproportionate increase in muscle mass with increasing body size ,17,20–22.kg-1.min-1or ml/min has the same limitations mentioned previously for ratio scaling (ml.kg-1.min-1) or absolute EE (ml/min). Also, as noted previously, as children age, a disproportionate change between RMR and EE of activity EE occurs [Some studies have used net EE to express the EE of an activity or changes in efficiency with age ,25,26. EE occurs ,28..kg-1.min-1 or 1 kcal.kg-1.h-1) is unacceptable for youth because the resting value used for the ADULT-MET (3.5 ml.kg-1.min-1) is typically less than the resting values for children (4–7 ml.kg-1.min-1). In addition, the ADULT-MET does not account for the physical changes that occur as children age, such as muscle mass to total mass ratio, leg length to stature ratio, and pubertal changes [y). However, Harrell et al. have shown that METy values are somewhat sex-dependent, tend to increase with age for low-intensity activities and decrease for vigorous-intensity activities, and are influenced by the weight and size of an implement being used for the activity .Another common approach to estimate EE in youth has been to describe the energy cost in terms of metabolic equivalents (MET) ,5,15,27. changes  and, henNormalizing youth EE to account for differences in growth and development could allow for comparisons of physical activities across ages and sex. Potentially, this might enable researchers to represent the energy cost of a physical activity using a single metric, regardless of the age of the youth, which could be useful for improving the Compendium of Physical Activities for Youth. However, no single study has compared the utility of these normalization approaches across a wide range of activities, in a heterogeneous sample of youth. Therefore, the goal of this study was to compare various approaches to normalizing the EE of children and adolescents for activities ranging from sedentary to vigorous.The initial sample of 583 boys and 477 girls  were obtained from five sites: Baylor College of Medicine , Michiga2 which was averaged per minute. The UM-B measured VO2 during the activities for eight minutes, while UNC measured the VO2 of each activity for 7 minutes. Further details on the site-specific instrumentation and protocols are available in the original publications . For this study all data were obtained under conditions designed to be at steady state as defined by each site. The oxygen uptake data were compared between the five sites for each activity and the data were found to be consistent across sites. Since walking and running protocols differed between sites these data were subject to further analyses as described below in the ‘Data Pooling and Cleaning’ section.Each site measured oxygen uptake, although the methodology differed. Baylor College of Medicine used a room respiration calorimeter with one-minute averaging and activities were completed for 10 to 20 minutes depending upon the intensity of the activity. Michigan State University and Oregon State University used calibrated OXYCON breath-by-breath, oxygen uptake systems with one-minute averaging and each activity performed for five minutes with rest between activities. Universities of Massachusetts–Boston (UM-B), and North Carolina (UNC) used calibrated COSMED portable metabolic units to measure breath-by-breath VO2.min-1) data to the first author. When available, measured resting VO2 also was submitted. Individual speeds for walking and running were reported and these speeds were grouped in 0.5 mph (0.8 kph) increments. Body mass index (BMI: kg.m-2) was computed from measured height (m) and body mass (kg) and BMI percentiles were obtained from CDC growth charts . In addition, we estimated RMR using the Schofield prediction equation [All study sites submitted their participant characteristics  and activity-related oxygen uptake (ml Oequation  because 2 (ml.min-1) served as the basis for calculating all other metrics. Two metrics were scaled for body mass (kg): VO2KG (ml.kg-1.min-1) and VO2ALLOM allometrically scaled to body mass (VO2.kg-0.75.min-1). Rather than trying to determine an allometric coefficient for each activity that could have varied by activity [2 for body surface area (ml.m-2.min-1) and height (ml.cm-1.min-1) because neither has been reported for more than a decade [2NET was estimated as VO2 of the activity in ml.kg-1.min-1, minus the child’s RMR (converted to VO2). Two additional metrics were also computed: YOUTH-MET (METy) was estimated as VO2 of the activity in ml.kg-1.min-1 divided by the youth’s estimated RMR (converted to VO2) and the ADULT-MET (VO2 in ml.kg-1.min-1 of the activity divided by 3.5 ml.kg-1.min-1) also was calculated [2KG, VO2ALLOM, VO2NET, YOUTH-MET, and ADULT-MET.Absolute VOactivity , we chosactivity ,22. We da decade . VO2NET lculated ,5. The f> 100 participants that included an age range of at least 8 to 18 years. We also used general linear models to determine whether the use of pooled data was acceptable, meaning the associations between oxygen uptake and age were similar across sites.The statistical analyses for this study comprised three phases–data pooling and cleaning, preliminary analyses, and final analyses. For data pooling and cleaning, we used descriptive statistics , t-tests and chi-square analyses were used to assess statistical significance of bivariate association, and graphical procedures  to 1) obtain a general picture of the sample, 2) investigate the distributional properties of the metrics, and 3) ensure each sedentary and aerobic activity was uniquely specific and had 2 for treadmill-walking at a speed of 2.5 mph (4 kph) and running at 4.5 mph (7.2 kph). All five sites reported VO2 for walking speed ranging from 2.8 to 3.2 mph (4.5 to 5.1 kph), some on treadmills and other self-paced. This 2.8 to 3.2 mph range will be referred to as “Walk 3 mph”. All four sites also reported self-paced or treadmill running at approximately 5 mph or 8 kph  in sufficient samples of children that we included this as a separate activity in our analysis . We excluded other sedentary activities, some of which combined several different behaviors  because the inclusion of multiple sedentary activities could result in a bias toward a metric that was most appropriate for low-intensity activities. We also excluded activities  deemed more anaerobic in nature. We examined the walking and running data and included only those youth who had specific speed data. Two sites reported the VOanalysis . Sensitianalysis . BasketbWe conducted preliminary analyses to identify a reduced set of metrics (from the five candidate metrics) that were least influenced by the physical characteristics of the children and therefore warranted a more comprehensive investigation. For the preliminary analyses, we calculated the correlation between the metric and age in years for each of the 14 activities, for each candidate metric. The correlations within each metric were averaged across activities (using Fisher’s Z transformation) , and theThe final analyses were conducted in three steps to: 1) identify the metric that was least associated with age, 2) determine whether the preferred metric was associated with the sex of the child, and 3) visually confirm the degree of variability by age within the reduced set of metrics. In doing so, we conducted a series of linear regression models with each candidate EE metric as the dependent variable and age as the independent variable. The standardized regression coefficient  measured the magnitude of association between each candidate EE metric and age–for the 14 activities. To test for differences in each of the standardized regression coefficients, we used the chi-square test for heterogeneity of a set of correlations ,31. For To examine whether the association between age and each EE metric varied by sex, we included age and sex as main effect terms and the age-by-sex interaction term in the regression models. A significant interaction term indicated the association between age and the EE metric was different for boys and for girls. A significant sex main effect term indicated that, after controlling for age, the EE metric for boys and girls was different. Finally, we used box-plots to visually confirm the variability and age-dependence of the optimal EE metric(s) for the 14 activities. Due to multiple testing, the global level of significance was set to p <0.01. Using Bonferroni’s adjustment, the level of significance for post hoc pairwise comparisons was set to p< 0.003. Data analyses were conducted with a statistical software package . All data are available as th percentile), 15.0% overweight (BMI 85th- 95th percentile) and 17.8% obese (BMI ≥95th percentile). The proportions in the three BMI categories were similar for both sexes. Overall, age was correlated with height  and body mass , but correlations were lower with BMI .The number of youth in each age group and their characteristics are presented by sex in 2 for the 14 physical activities are displayed in y values for all activities by an average of 0.9±1.2 METs.The means (±SD) for the metrics of VO2NET, VO2ALLOM, YOUTH-MET, VO2KG, and ADULT-MET, respectively. Figs 2, VO2KG, VO2NET, VO2ALLOM, and METy for six representative activities of various intensities. The reader should keep in mind that the optimal metric should have slopes that remain horizontal with increasing age. The figures point out that absolute VO2 increases with age for all activities  with the least age-dependency (weakest correlations) were further examined in the final analysis.The rank-ordered (weakest to strongest) averaged correlations for each metric with age (using Fisher’s Z transformation) were -0.11, -0.23, 0.28, -0.37, and -0.38 for VOtivities , while Vsketball . Fig 3 s scaling  results   are presented in y was not statistically influenced by age, except for television viewing, in which case the correlation was low (r = 0.149). For non-weight bearing cycling, VO2NET and VO2ALLOM were most independent of age; METy was positively correlated with age (r = 0.535). For moderate-to-vigorous activities, VO2ALLOM generated the lowest correlations and thus, was least influenced by age. For activities requiring more motor skills , the results were inconsistent.The standardized regression (correlation) coefficients between age and EE based on the three remaining metrics .Results from the activity-specific chi-square test for multiple correlations showed the correlation for at least one metric was significantly different than the other two metrics for all activities (p < 0.001). 2ALLOM (R2 = 0.051), VO2NET (R2 = 0.069), or METy (R2 = 0.019).Follow-up analyses revealed that for most activities, the age-by-sex interaction (PROC GLM) was not significant (p > 0.001), indicating that in the majority of cases, that association between the EE metrics and age did not differ by sex. The main effect for sex was not consistently significant; there was only minor evidence of effect modification by sex for the activity of rope skipping (p < 0.05) normalized as VOy is independent of age and sex. For moderate-to-vigorous activities, VO2ALLOM was least influenced by age, but most influenced by sex (p <0.0001). For the more skilled activities the findings were inconsistent, probably related to differences in performance capabilities, experience, or exposure that occurs with age.Of the EE metrics examined, our findings suggest that no metric completely eliminated the influence of age, height, body mass, or sex for all 14 activities. For most sedentary and light-intensity activities, it appears that METThe preferred metric for standardizing EE of children is dependent on the type and intensity of the physical activity. Some of the activities were controlled ; whereas others were performed at self-selected intensities. These volitional activities would be expected to have greater variation in EE among children because of differences in: 1) the fitness level of the child , 2) the 2NET or METy , 7.0 for a boy age 11 years (RMR = 4.3 ml/kg/min), and 8.8 for a boy age 16 years (RMR = 3.5 ml/kg/min). Overall, the use of the METy has limitations for moderate-to-vigorous activities because of the influence of age. The METy does have an advantage, however, over the other metrics with respect to eliminating any differences related to the sex of the child. The differences between the sexes for all activities were < 0.7 METy.Our findings showed that the METproposed ,15,27; h.kg-1.min-1 to compare the energy cost of treadmill exercise in boys and men [2 ~33%) between the EE expressed per kilogram body mass and the child’s physical characteristics, including age (Early studies by Robinson used oxygen uptake expressed as the metric of ml and men . Our resding age . This ma2NET) might stabilize the energy cost with respect to the differing physical characteristics of the children. Our results show that this was not the case  has been used to estimate energy expenditure in youth [.kg.-1min-1) is typically less than the resting values for children, that can range from 4 to 7 ml.kg.-1min-1. Our results show that applying the ADULT-MET value overestimates the METy values of most activities by at least one METy, which would lead to an over-estimation of the EE of the activity. In addition, the ADULT-MET does not take into consideration the physical changes that occur as children age [The ADULT-MET . We did not evaluate the impact of BMI status on the age-dependency of the EE metrics . Howeverpendiums ,5, will y  displayed the least age dependence for sedentary and low-intensity activities and it was not influenced by the sex of the child. VO2ALLOM was the best for reducing age-dependence for moderate-to-vigorous intensity activities. Although VO2ALLOM appears to work well for ambulatory activities, it did not work as well for activities that require additional equipment or more skills such as playing basketball. VO2ALLOM coefficients are complex to calculate and a different mass exponent may be required for each activity, making it challenging to directly compare activities across studies. In addition, some target audiences may have difficulty interpreting and utilizing these measures.A common metric to express energy expenditure for sedentary behaviors and physical activities in children and adolescents is needed to evaluate interventions and policies to increase youth physical activity. Our findings show it is difficult to clearly identify a single metric to accurately express the energy expenditure of common activities across a wide age range; thus, there is no optimal metric. In our study, none of the EE metrics we examined completely adjusted for differences in the age, sex, and physical characteristics of children and adolescents. Of the metrics we examined, the MET2ALLOM or METy values is important to consider. These issues are important for the further development and improvement of a Compendium of Physical Activities for Youth. Based on our findings and these considerations, we would recommend the use of the METy recognizing and accounting for its age-dependency, especially for the more vigorous activities. Age is a key factor related to energy expenditure for youth both at rest and during physical activities and should be accounted for in developing common metrics for various physical activities in children and adolescents.The optimal metric for expressing EE may depend on at least two additional issues not addressed in this study–translation and generalizability. The ability of specific target audiences to calculate, interpret, and utilize VOS1 Data(PDF)Click here for additional data file.


The current study investigated the time course of the other-race classification advantage (ORCA) in the subordinate classification of normally configured faces and distorted faces by race. Slightly distorting the face configuration delayed the categorization of own-race faces and had no conspicuous effects on other-race faces. The N170 was sensitive neither to configural distortions nor to faces' races. The P3 was enhanced for other-race than own-race faces and reduced by configural manipulation only for own-race faces. We suggest that the source of ORCA is the configural analysis applied by default while processing own-race faces. It is documented that although humans can identify faces of their own race better than of other-race faces  as well as featural processing and hence, the role of the second-order configural processing in faces' race detection is not reliably revealed by N170 although it is not sensitive to second-order configural processing . All participants had normal or corrected to normal visual acuity and had no history of psychiatric or neurological disorders. They were right handed based on self report and were paid for participation. All participants signed an informed consent approved by the Ethical Committee of General Hospital of People's Liberation Army and were paid for their participation. One subject was excluded from the analysis due to artifacts during EEG recording.M = 2.3, SD = 0.5, indicating the participants had relatively little experience with faces from the other race.Either before or after the electroencephalogram (EEG) recording , participants completed one self-report questionnaire regarding other-race contact, including five items modeled after Walker et al. . Item (iwww.adobe.com) .Stimuli consisted of 70 grayscale photographs of Chinese  and 70 Caucasian . All faces were of young people (20–30 years old) and were unfamiliar to the participants. The distortion of second order relations in each of these faces were made by reducing the distance between the eyes by 20%, lowering the eyes' level relative to the tip of the nose by 20% and reducing the distance between the root of the nose and the mouth by 20%. All images were equated for luminance and root mean square (RMS) contrast  and the emotional expression in stimuli was controlled , using Adobe Photoshop  Figure . IncludiThe 280 stimuli were randomized and presented at the center of a computer monitor in two blocks of 140 stimuli each separated by a short break. Each stimulus was exposed for 500 ms and followed by an inter-stimulus interval (ISI) that varied randomly between 2000 and 2500 ms. Participants were instructed to classify each stimulus by race (Chinese or Caucasian) as quickly and as accurately as possible and report their decision by pressing alternative buttons one with the index finger of the right-hand and the other with the index finger of the left hand. The assignment of the buttons to response alternatives was counterbalanced across participants.The experiment was run in an electrically isolated and sound-attenuated booth. Following electrode montage and instructions a short training set including 12 normal configured faces and 12 distorted faces equally from each race were presented using the same procedure as in the actual experiment. None of the training stimuli were used in the experiment.www.ant-neuro.com). The band-pass filter range of 0.1–100 Hz was used during EEG recording at both locations. Vertical EOG and horizontal EOG were recorded with two pairs of electrodes, one placed above and below right eyes, and another 10 mm from the lateral canthi. An additional electrode as the reference electrode was placed on the tip of the nose. Throughout the EEG recording, the impedance of the electrodes was kept under 5 kΩ.The electroencephalogram (EEG) was recorded by 64 Ag/AgCl electrodes cap according to the extended 10/20 system and continuously sampled at 512 Hz by ANT asaLab (www.ant-neuro.com). After EOG artifacts correction, remaining artifacts exceeding ±100 μV in amplitude or containing a change of over 100 μV within a period of 50 ms were rejected at both locations. Artifact-free EEG was then segmented into epochs ranging from 200 ms (as the baseline correction) before to 800 ms after stimulus onset and averaged separately for each participant and for each of the 4 conditions Chinese normal faces, Chinese distorted faces, Caucasian normal faces, and Caucasian distorted faces. Only correct responses were included in the average but all averages included at least 50 trials. The averaged waveforms were digitally low-pass filtered at 30 Hz (24 dB/octave) to reduce high-frequency noise.EEG data processing was performed off-line using ASA 4.9 (Accuracy rates and RTs (from the stimulus onset) were recorded and analyzed with Race-of-the-stimulus  and Configuration  as the within-subjects factors.In line with most previous studies of N170  = 71.2, MSE = 85027, p < 0.001; partial η2 = 0.59] and Configuration , reflecting that, overall, other-race faces were identified more quickly (525 ± 90 ms) than own-race faces (565 ± 83 ms), i.e., ORCA, and altering configuration of faces delayed race classification . Importantly, the two-way interaction of Race-of-the-stimulus * Configuration . The post-hoc analysis of this two-way interaction revealed that the ORCA was significant for both normally configured faces  and for distorted faces , but it was significantly larger for the latter than for the former faces set (p = 0.008 < 0.01), and that the changes of facial configuration delayed race classification only for own-race faces (p = 0.007 < 0.01) not for other-race faces (p > 0.1).For each participant and experimental condition RTs that were more extreme than ±2 SD from the mean have been excluded (less than 2%). The ANOVA revealed significant main effects of Race-of-the-stimulus , i.e., the ORCA of the accuracy. The main effect of Configuration was significant  but qualified by the Race-of-the-stimulus * Configuration interaction . Further post-hoc analyses showed that the ORCA of accuracy was significant for distorted faces  but not for normal configured faces  and that the configuration modulation reduced the accuracy for classifying own-race faces (p = 0.007 < 0.01) but not for classifying other-race faces (p > 0.05).For the analysis of accuracy, the other-race faces were identified more accurately (95 ± 5%) than own-race faces , and larger at PO7/8(−11.6 ± 2.6 μV) than at P7/8 (−10.7 ± 2.8 μV), and largest at P9/10 (−12.3 ± 3.0 μV) .Neither the amplitude nor the latency of the N170 was affected by the Race-of-the-stimulus (F < 1.0), but significant main effect of Configuration was found  and this effect was qualified by the significant interaction of Race-of-the-stimulus × Configuration . Post-hoc analysis showed that distorting face's configuration enhanced N250 to Caucasian (other-race) faces  but no effect on Chinese (own-race) faces . The main effect of Electrode site  and interaction of Electrode site × Hemisphere  were also found, reflecting significant main effect of Electrode site at left hemisphere  but not at right hemisphere . No other main effects and interactions were found.The mean amplitudes of N250 component (200 ~ 300 ms post stimuli onset) showed no main effect of Race-of-the-stimulus  = 42.4, MSE = 929, p < 0.001; partial η2 = 0.46], that is the other-race advantage (ORA) of P3 (ORA-of-P3), and that configural distorted faces elicited smaller P3 amplitudes (11.8 ± 2.2 μV) than normally configured faces . We also found a marginally significant interaction of Configuration * Race-of-the-stimulus  and post-hoc analysis revealed that whereas for own-race faces the amplitude of the P3 was larger for normally configured (12.2 ± 2.8 μV) than distorted faces , no such effect was found for other-race faces  and that the ORA-of-P3 was larger for distorted (3.8 ± 1.8 μV) than normal faces . The Site effect was also significant .ANOVA confirmed that the P3 amplitude was higher for other-race faces (13.4 ± 3.2 μV) than own-race faces , which suggested that whereas for own-race faces the P3 latency for distorted faces was shorter (482 ± 48 ms) than for normally configured faces , there was no such difference for other-race faces . Note that the measurement of the P3 latency elicited by distorted own-race faces might also reflect a negative-going potential that interrupted the P3  not between the RTs and the amplitude of N170 component (p > 0.1), that is, the longer the RTs the lower the P3 amplitude. The correlations between the RTs and the latencies of ERP components were not found (ps > 0.1).In addition to the ANOVAs, we calculated the Person correlations between the RTs and the peak latencies/amplitudes of the N170 and P3 components. Significant negative correlations were found between the RTs and the amplitude of P3  = 18.2, MSE = 507, p = 0.007 < 0.01]. The main effect of Site was also significant , reflecting the maximum of Pd3 at Cz site (3.2 ± 1.8 μV), i.e., central-parietal distribution as presented in Figure For Pd3 component, mean amplitude between 450 and 550 ms was analyzed by a Two-way ANOVA with Configuration  and Site  as within-subjects factors. ANOVA showed that larger Pd3 was found for distorted faces (3.8 ± 1.6 μV) than original faces [1.5 ± 0.8 μV, F = 18.2, MSE = 508, p < 0.001; partial η2 = 0.3.For CNd component, mean amplitude between 450 and 550 ms was analyzed by a Two-way ANOVA with Race-of-the-stimulus  and Site  as within-subjects factors. The ANOVA showed that the CNd component was more negative for own-race (−2.4±0.8 μV) than other-race faces (−0.2 ± 0.9 μV), In the current experiment we investigated the involvement of configural computations while processing own-race and other-race faces in a race categorization task. Analyzing performance, we found that in Chinese participants the classification of other-race faces was faster than the classification of own-race faces (ORCA). Importantly, however, the ORCA on the classification speed was twice as big for distorted faces than for normally configured faces and the second-order configural modulation reduced the performance for own-race faces not for other-race faces. Because the configural computation was task-irrelevant in the present study, these data indicated that one of the sources of ORCA is the configural analysis applied by default while processing own-race faces, replicating our previous behavioral study  and the plasticity of an acquired familiarity that accrues gradually over time, respectively . In line with recent findings using the same race classification task  faces, the configural effect was indeed appeared the earlier time window (200 ~ 400 ms) mainly relevant to the deflection of the N250 component. One possible explanation for configural effect for other-race faces is that participants may detect the configural changes of other-race faces at earlier stage of face identification by semantic and/or individual information  might reflect a negative-going potential  that interrupted the P3 (see Figure In sum, although Chinese participants were tested only in the present study, along with the similar interaction found for RTs, the fact that the effect of configuration of P3 was significant for own-race faces but not for other-race faces implicates that the ORCA is caused by the default tendency to process own-race faces to the individual level whereas other race faces are initially perceived at the race-group level, at least for Chinese participants. In other word, during the explicit face classification by race, other race faces are perceived as a group, whereas the grouping of own-race faces is interfered with and delayed by the automatic application of configural computations that are necessary for distinguishing among individual faces but probably irrelevant for race categorization. Due to the fact that Chinese participants were recruited only in the present study, a cross-culture research, i.e., two race groups of observers at least, would be necessary to clarify the present conclusion in the future.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The Review Editor Lucy Troup declares that, despite being affiliated with the same institution as the Associate Editor Carol Seger, the review process was handled objectively.


Human T-lymphotropic virus type 1 (HTLV-1), a retrovirus, is the causative agent of HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) and adult T-cell leukaemia/lymphoma (ATLL). The reported association with pulmonary disease such as bronchiectasis is less certain.A retrospective case review of a HTLV-1 seropositive cohort attending a national referral centre. The cohort was categorised into HTLV-1 symptomatic patients (SPs) ) and HTLV-1 asymptomatic carriers (ACs). The cohort was reviewed for diagnosis of bronchiectasis.34/246 ACs and 30/167 SPs had been investigated for respiratory symptoms by computer tomography (CT) with productive cough +/- recurrent chest infections the predominant indications. Bronchiectasis was diagnosed in one AC (1/246) and 13 SPs   with high resolution CT. In the multivariate analysis ethnicity (p = 0.02) and disease state (p < 0.001) were independent predictors for bronchiectasis. The relative risk of bronchiectasis in SPs was 19.2  and in HAM/TSP patients compared with all other categories 8.4 . Subjects not of African/Afro-Caribbean ethnicity had an increased prevalence of bronchiectasis .Bronchiectasis was common in the cohort (3.4 %). Risk factors were a prior diagnosis of HAM/TSP and ethnicity but not HTLV-1 viral load, age and gender. The spectrum of HTLV-associated disease should now include bronchiectasis and HTLV serology should be considered in patients with unexplained bronchiectasis. The spectrum of disease associated with HTLV-1 has not been completely defined. The association between HTLV-1 and pulmonary conditions is less well documented and clinically recognised. Bronchiectasis has been reported amongst Australian aborigines infected with HTLV-1. We have observed a high rate of bronchiectasis in a UK HTLV-1 infected cohort. We have shown that patients with non-pulmonary HTLV-1-associated disease, particularly inflammatory disease, are more likely to have an additional diagnosis of bronchiectasis than asymptomatic carriers and postulate that the same dysregulated hyper-inflammatory response seen in HTLV-associated myelopathy/tropical spastic paraparesis may play a role in this setting. Our findings highlight the importance of screening HTLV-1 infected patients, particularly those with inflammatory disease, for pulmonary disease.The human T-lymphotropic virus type-1 (HTLV-1), a retrovirus, has been estimated to infect at least 5-10 million people worldwide with the highest prevalence reported in Japan, Africa, The Caribbean and South America , with a Although less recognised associations between HTLV-1 and pulmonary conditions have been reported: patients with HTLV-1 have increased lymphocytes in bronchoalveolar lavage fluid (BAL) which correlates with HTLV-1 VL in both the BAL and peripheral blood mononuclear cells (PBMC) ; patientet al. [To date the most comprehensive report of pulmonary imaging in HTLV-1 infection is that of Okado et al.  who undeet al. . Two addet al. , 27. A cet al. . Furtheret al. . A causaet al. . It is tAlthough HTLV-1 associated diseases are rare in the UK the National Centre for Human Retrovirology manages a relatively large HTLV cohort, the only one of its kind in Europe. The cohort was initially established in 1993 to gain an understanding of the breadth and pathogenesis of HTLV disease through longitudinal observation. Despite the aforementioned reports HTLV-1 infected patients are not generally considered to be at greater risk of bronchiectasis than the general population and therefore chest imaging in our cohort has only been conducted if there was a clinical suspicion of lung disease.A single national centre retrospective case review of all 464 HTLV-1 seropositive patients attending the National Centre for Human Retrovirology, St Mary’s Hospital, London, UK between 1993 and 2012. Subjects were characterised in regard to HTLV-1 infection as either symptomatic patients (SP) or as ACs. SPs had an underlying disease with documented association with HTLV-1: ATLL; strongyloidiasis; HAM/TSP; or other HTLV associated inflammatory disease (HAID) namely polymyositis, uveitis and arthritis. Twenty-three patients were excluded as they were co-infected with human immunodeficiency virus type 1. A further 28 patients were excluded as they had neurological features that could not be classified as either HAM/TSP or polymyositis and it was difficult to establish whether HTLV-1 infection was causal or co-incidental and thus whether they had underlying HTLV-1-associated disease.The clinic database, case notes, laboratory and radiology reports were examined. In accordance with the guidance produced by the UK National Research Ethics Service, research ethics approval was not required for this study. Once the data were collated we used the clinical code, which is separate from the hospital number, to ensure confidentiality. Bronchiectasis was correlated with HTLV-1 symptoms and with concurrent HTLV-1 VL (this being the number of HTLV-1 DNA copies/100 PBMCs and reported as VL %). Data were collated on the number of patients that had CT i.e. CT chest or high resolution CT (HRCT), the clinical indication for the imaging and the radiological findings particularly the presence or absence of bronchiectasis. A multi-detector CT scanner was used. The CT criteria that were used for diagnosis of bronchiectasis were bronchial dilatation as evidence by bronchoarterial ratio greater than 1 to 1.5 (signet ring sign) and/or non-tapering airways . Indirect signs such as thickening of the bronchial wall, loss of lobe volume, mosaic perfusion pattern, a tree-in- bud pattern, and mucus plugs were also used.t-test, relative risk (RR), Fisher’s exact test and backwards multivariate logistic regression using commercial statistics software . A p-value of 0.05 or less was considered significant.Data was also collected on risk factors for bronchiectasis i.e. congenital conditions, tuberculosis, childhood infection and exposure to chemical irritant. We also collected data on the presence of other chronic conditions i.e. chronic liver disease, myocardial disease, renal and cardiac failure. Data were analysed by Student Four hundred and thirteen patients infected with HTLV-1 were included comprising 246 ACs and 167 SPs. Of the 167 SPs: 54 had ATLL; 95 HAM/TSP; 11 HAID and 7 strongyloidiasis , SPs were significantly more likely to be of African/Afro-Caribbean ethnicity  than the ACs   compared with ACs . HTLV-1 VL among non-HAM/TSP SPs was higher still . The median VLs in the different SP groups were: HAM/TSP 14 %, HAID 8.5 %, ATLL 29.3 % and Strongyloidiasis 20.6 %  in the AC group and 10/30 (33.3 %) in the SP group were high resolution CTs. (HRCTs). The reasons for CT imaging were productive cough +/- recurrent infection (50 % ACs vs. 50 % SPs), weight loss (26.5 % ACs vs. 23.3 % SPs), suspected malignancy (23.5 % ACs vs. 6.7 % SPs) or an abnormal chest X-ray (SPs 16.7 %). The indication for CT was unclear in 3.3 % SPs. The indications for HRCT was productive cough +/- recurrent infection. The time from HTLV-1 diagnosis to imaging was on average 2.5 years in ACs and 2 years in SPs  and ethnicity (p = 0.02) but not age, gender or HTLV-1 VL were independent predictors for bronchiectasis.Our data suggest that bronchiectasis is more common among HTLV-1 infected subjects in the UK than among the general population, that the inflammatory process of HAM/TSP and other HAID is implicated in pathogenesis and that Caucasians infected with HTLV-1 may be more susceptible to bronchiectasis than African/Afro-Caribbeans.Mass chest radiography during the 1950s indicated a prevalence of 100 per 100,000 adult UK population of bronchiectasis . The 3.4In the general population the prevalence of bronchiectasis increases with age. A retrospective cohort study in the US found the prevalence of bronchiectasis to be 4.2 per 100,000 population aged 18-34 years and 271.8 per 100,000 population aged >75 . HoweverAnother consideration is the ethnic bias of the cohort which is predominantly of African/Afro-Caribbean ethnicity. However not being African/Afro-Caribbean carried a greater than 3 fold increased risk for diagnosed bronchiectasis. Cohort data from the two Caucasian populations with highest HTLV-1 prevalence, Iran and Romania have not been reported. Whilst the lower risk of bronchiectasis observed in African/Afro-Caribbean subjects merits further investigation this may account for the paucity reports of association between HTLV-1 and bronchiectasis from outside of Japan. Genetics, timing of primary infection and environmental factors may all contribute to these different rates. Difference in bronchiectasis rates amongst different ethnicities have been reported from New Zealand and Fiji between Polynesian and Non-Polynesian and Melanesian and Indian children respectively , 33. WhiWe hypothesised that HTLV-1 VL would be associated with bronchiectasis and as previously reported for this cohort, and consistent with other studies, HTLV-1 VL was significantly higher in patients with symptomatic HTLV-1 infection than in the ACs . HoweverOur HTLV-1 cohort is comprised of patients from the three main categories of infection, namely asymptomatic carriage, ATLL and HAM/TSP and, as reflects the demographics of HTLV-1 infection in the UK, was predominantly of African/Afro-Caribbean ethnicity. The high number of patients that were Caucasian in the AC group is secondary to universal blood donor screening. The M: F ratio between ACs and SPs was similar. A major cause for bronchiectasis is early childhood infections. This can be difficult to exclude however there was no significant childhood infections reported by the patient or in their health records. Further to this even though we did not test for autoimmune conditions or alpha 1 antitrypsin deficiency our patients had no clinical symptoms or signs of these conditions.Strongyloides stercoralis, with a higher incidence HTLV-1 amongst S. stercoralis carriers [The pathogenesis of bronchiectasis associated with HTLV-1 has not been studied. The association here with a prior diagnosis of another inflammatory disease and the prominent lymphocytosis in BAL reported by others , 21 poincarriers , and a hcarriers .The major limitation of this study is is that CT imaging had only been performed if the patient had clinical or radiological features that raised the suspicion of underlying pulmonary disease with no lower threshold for imaging in this cohort compared to the general population. Thus only 15 % of the cohort had CT imaging (13.8 % ACs and 18.0 % SPs) and the prevalence of bronchiectasis, particularly early/mild or asymptomatic, may have been underestimated. Second, the number of cases of bronchiectasis is relatively few. Despite this the associations reported are statistically robust, suggesting that these are not chance findings. Third, we have no data on the effect of HTLV-1 or associate inflammatory cells in the lung and can only speculate regarding pathogenesis. The relative strengths are studying a population that is socially and ethnically distinct and the median seven years follow-up of each patient group. A prospective study is now justified to confirm and extend these findings on the frequency of bronchiectasis in patients with HTLV particularly amongst those with pre-existing HTLV-associated disease and to explore pathogenesis.In the UK bronchiectasis is more common amongst HTLV-1 infected subjects (3.4 %) than reported for the general population (0.1 %). Non-African/Afro-Caribbean patients with pre-existing HTLV-1-associated disease are most at risk. Our data support the view that the spectrum of HTLV-1-associated disease is broader than previously thought and that disseminated mild inflammation may be common, albeit asymptomatic. Our results emphasise the importance of screening symptomatic HTLV-1 patients for pulmonary disease  early following diagnosis of HTLV-1 and that CT imaging should be requested even in the presence of minimal symptoms. Furthermore HTLV serology should be considered in all patients with bronchiectasis in whom no other cause is apparent.